Cholecystokinin (CCK) is produced by discrete enteric endocrine cells (I cells) of the proximal small intestine and secreted into the blood upon ingestion of food. However, the exact nature by which the CCK cell is regulated is unknown. Recently, the PI has developed a method for studying CCK cells in vitro. These cells respond to a variety of physiological stimuli including the novel CCK releasing factors, monitor peptide and luminal CCK releasing factor (LCRF). Although these cells can be enriched they are not a pure population of enteric endocrine cells. Therefore, the PI has recently characterized an intestinal cell line (STC -1) that produces high amounts of CCK and exhibits properties characteristic of native enteric CCK cells. This established cell line secretes CCK in a regulated manner that is indistinguishable from native CCK cells and has been used as a model for studying electrophysiological properties of intestinal hormone secretion. In other endocrine systems hormone secretion is intimately coupled to plasma membrane ion channel activity. The PI has previously demonstrated that maneuvers that cause membrane depolarization in STC-1 cells induce CCK secretion. Using patch clamp recording techniques it has been shown that CCK-secreting cells have a prominent basal potassium current. Moreover, calcium channel activity appears to be critical for CCK release. These findings suggest that regulation of ion channel activity in CCK cells may be important for regulated CCK secretion. The hypothesis to be tested in the current proposal is that CCK releasing factors regulate plasma membrane ion channel activity in intestinal endocrine cells to stimulate hormone secretion. For these studies, ion channel activity will be measured using patch clamp recording techniques and membrane potential will be measured with conventional microelectrodes. The following specific aims will be addressed: 1. To determine whether CCK secretion induced by monitor peptide and LCRF occurs through changes in membrane depolarization or by direct effects on ion channels. 2. To characterize the specific potassium channels which are regulated hi CCK cells by releasing factors. 3. To identify the calcium channels that are regulated by monitor peptide and LCRF. Together these studies will determine how intestinal releasing factors regulate CCK secretion. state)